Accepted Manuscript

Title: Design and study of poloxamer-based microemulsion

gels with naproxen

Authors: Anna Froelich, Tomasz Osmałek, Paweł Kunstman,

Barbara Jadach, Monika Brzostowska, Wojciech Białas

PII: S0927-7757(18)31065-3
DOI: https://doi.org/10.1016/j.colsurfa.2018.11.006
Reference: COLSUA 22976

To appear in: Colloids and Surfaces A: Physicochem. Eng. Aspects

Received date: 13 September 2018

Revised date: 24 October 2018
Accepted date: 2 November 2018

Please cite this article as: Froelich A, Osmałek T, Kunstman P, Jadach B, Brzostowska
M, Białas W, Design and study of poloxamer-based microemulsion gels with
naproxen, Colloids and Surfaces A: Physicochemical and Engineering Aspects (2018),
https://doi.org/10.1016/j.colsurfa.2018.11.006

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Design and study of poloxamer-based microemulsion gels with naproxen

Anna Froelich1*, Tomasz Osmałek1, Paweł Kunstman1, Barbara Jadach1, Monika

Brzostowska1, Wojciech Białas2

1
Poznan University of Medical Sciences, Department of Pharmaceutical Technology, ul.
Grunwaldzka 6, 60-780 Poznań, Poland, tel. +48 61 854 66 61

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2
Poznan University of Life Sciences, Department of Biotechnology and Food Microbiology,
ul. Wojska Polskiego 48, 60-627 Poznań, Poland

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*
[email protected]

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Graphical abstract

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Abstract
In this study novel microemulsion-based gels with poloxamer 182 as a surfactant were
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designed and investigated as potential carriers for topical dosage forms with naproxen, non-
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steroidal anti-inflammatory drug. Based on the analysis of the pseudoternary phase diagrams
obtained with the use of different components, oleic acid was selected as an oil phase and
ethanol, isopropanol and Transcutol® were used as cosurfactants. As a polar phase, pure
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water and poloxamer 407 solutions were tested. The extent of monophasic areas was similar
in both cases, however, the poloxamer solutions were excluded from further experiments
because of the lack of gelling properties in the studied systems. As a thickening agent,
Carbopol® EZ-3 was selected. In three microemulsion systems with different cosurfactans
electrical conductivity was analysed as a function of polar phase content in order to find the

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transition points corresponding to microstructural changes leading to different microemulsion
types. In the next step, three bicontinuous systems were transformed into semisolid polymer
gels and subjected to rheological and texture profile analyses in order to assess mechanical
parameters important in terms of topical drug delivery.

Keywords
microemulsion, naproxen, gel, rheology, texture profile analysis

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1. Introduction

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Microemulsions have been a subject of numerous studies concerning their

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structure, physicochemical properties and potential application in many different
industrial and medical areas [1–4]. Microemulsions are nanodispersive systems with a

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diameter of droplets usually varying from 10 to 100 nm [5]. These values do not exceed
the visible light wavelength which results in ME transparency [6]. Another important feature

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of these systems is their thermodynamic stability and spontaneous formation process which is
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caused by extremely low interfacial tension between immiscible oil and water phases. In order
to decrease interfacial tension values usually a mixture of surfactant and cosurfactant is
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necessary [7]. One of the most extensively studied properties of microemulsions is their
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ability to undergo a microstructural transition known as a percolation. In this process,

the system transforms from one structural type into another, e.g. from w/o into a
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bicontinuous one or from a bicontinuous one into o/w. The mentioned transitions are
related either to the changing ratio between polar and nonpolar phase or to the
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temperature changes. The described structural changes are accompanied by the changes
in electrical conductivity of the system. Therefore, the analysis of the electrical
conductivity of microemulsion systems may be applied to assess the boundaries between
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the regions corresponding to particular microemulsion types.

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Because of the unique properties including ease of preparation, high stability and
excellent solubilizing properties, MEs are frequently investigated as potential new carriers in
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drug delivery. Apart from the mentioned physicochemical features important for
pharmaceutical technology and the design of the industrial process, it was also shown that
MEs have an enormous potential in improving the bioavailability of the active pharmaceutical
ingredient (API). This is particularly valuable in the case of the drugs revealing poor
solubility in water and poor permeability through biological membranes. It is noteworthy that
several commercial products classified as self-microemulsifying drug delivery systems

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(SMEDDS) have been successfully introduced to the pharmaceutical market [8]. These
products consist of oil, surfactant and cosurfactant and are intended for oral administration.
ME is formed upon the contact with the gastrointestinal fluid which acts as a water phase for
these systems. To the best of our knowledge, there are no ME-based marketed products
intended for other administration routes. Nevertheless, numerous studies show that they can
also improve biopharmaceutical characteristics of the topical and transdermal products [9–
11]. The interaction between skin and ME is explained by several different theories [12,13].
ME components such as a surfactant, medium- or short-chain alcohols and oils are skin

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absorption promoters which reversibly disrupt lipid layers in stratum corneum and change its

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permeability. Furthermore, in o/w MEs drugs are encapsulated in the inner phase which may

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act as a reservoir to maintain the constant gradient between the external phase and skin
surface [12]. However, despite all advantages revealed by MEs their low viscosity may appear

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as a problem leading to difficulties with the administration to the skin [14]. To minimize this
effect the thickening agent can be introduced into the system in order to transform it from
liquid to semisolid state.
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Poloxamers are triblock copolymers composed of centrally located poly(propylene
oxide) block (PPO) connected to two peripheral poly(ethylene oxide) blocks (PPO). PEO are
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more hydrophilic than PPO blocks which makes the whole molecule amphiphilic. It is
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important to notice that the properties of the polymer can be modulated by the changes in the
ratio between hydrophobic and hydrophilic parts of the molecule. In pharmaceutical
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technology poloxamers can be used as solubilizers [15], surfactants [16] and some of them,
e.g. poloxamer 407, as thickening agents [17].
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Semisolid gels are composed of an external phase immobilized with the three-dimensional
structure formed by a thickening agent. Depending on the polarity of the dispersing medium
they can be classified as hydro- or organogels. The most popular gelling agents employed in
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many different industrial areas are polymers, however, it is also possible to obtain semisolid
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systems with low molecular mass organogelators [18] and inorganic compounds [19]. In
topical and transdermal drug delivery gels are considered as attractive carriers for the active
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substance, displaying numerous advantages over the other semisolid and liquid dosage forms.
They are usually less greasy and can be washed easier than ointments. Moreover, they provide
better residence time at the application site and more convenient application than liquid
formulations. It was shown that selecting a proper vehicle for the topical product is
crucial for the patient adherence to the treatment regimen [20,21]. Low acceptability of
the product caused by the improper selection of the vehicle may result with treatment

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discontinuation. Mechanical properties of the topical formulation are important for the
application process and for perception and acceptance of the product. It is also important to
note that mechanical characteristic is essential for the manufacturing process including mixing
and filling the containers. In order to evaluate the properties important both in terms of
manufacturing and product application to the skin surface, several analytical techniques
have been applied. Rheological investigations are generally based on the observation of
the sample response to the shearing forces which also occur during the product
application or manufacturing. As a result, valuable practical data, e.g. information on

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shear-thinning or thickening behavior and viscoelastic properties, may be obtained.

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Another technique less frequently employed in the investigations concerning

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pharmaceutical formulations in texture profile analysis (TPA). Texture profile analysis
was initially developed and applied in food science and technology [22]. The first reports

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introducing this technique in the field of pharmaceutical analysis were published by
Jones et al. [23–26]. The available literature reports employing this method in the

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description of various dosage forms show that it may be used as a simple, fast and
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reliable complementary technique with respect to rheological studies [27]. The
parameters like hardness, cohesiveness, adhesiveness and compressibility obtained from
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texture profile provide valuable information on the behavior of the sample subjected to
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mechanical stress.
The experimental data may give an insight into the behavior of the investigated material
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in the technological processes involving the application of shear forces, e.g. mixing and
pumping into the containers. Moreover, whenever topical use of the semisolid product is
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taken into consideration the results of rheological and textural studies may be useful for
prediction of the material behavior during spreading on the skin. The application
aspects are important in terms of the patients' comfort and also for the residence time at
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the application site which related directly to the therapeutic effect.

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In this paper, we focused on the design and evaluation of microemulsion-based gels

employing poloxamer 182 as a surfactant. In the first step several systems with different
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compositions have been checked for the ability to form microemulsions and the systems
with highest area corresponding to transparent isotropic liquids in pseudoternary phase
diagrams were selected. In the selected systems electrical conductivity was investigated
as a function of water content in order to obtain information on the microemulsion
structure. The main objective of the presented study was to analyze in detail the impact of
the system composition on the rheological and textural properties of microemulsion-based

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polymer gels intended for topical drug delivery. As a model active substance naproxen, non-
steroidal anti-inflammatory drug (NSAID) was used and the effects related to its presence in
the semisolid systems were described.

2. Materials and methods

2.1.Materials
Ethanol 96% was purchased from Avantor Performance Materials Poland S.A.

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(Gliwice, Poland). Ethyl oleate (EO), oleic acid (OA), Kolliphor® P 407 and Transcutol® P

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were purchased from Sigma-Aldrich (Saint Louis, MO, USA), Isopropyl myristate (IPM),

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propylene glycol (PG) and isopropanol were purchased from P.P.H. "Stanlab" (Lublin,
Poland). Synperonic™ PE/L 62 was obtained free of charge from Croda Poland (Kraków,

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Poland). Carbopol® EZ-3 was a generous gift from Lubrizol (Wickliffe, OH, USA).
Naproxen (NPX) was provided as a free sample by EMO-FARM (Ksawerów, Poland). Water

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used for the experiments was purified and deionized by the Simplicity® Water Purification
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System (Merck Millipore, Billerica, MA).
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2.2.Construction of pseudoternary phase diagrams

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In order to find out the concentration range of components for the existence range of
microemulsions, pseudoternary phase diagrams were prepared with the use of polar phase
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titration method. As an oil phase EO, IPM and OA were used. Poloxamer 182 (Synperonic™
PE/L 62) was used as a surfactant (S), while ethanol, isopropanol, Transcutol® P and
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propylene glycol were investigated as cosurfactants (C). As a polar phase, deionized water,
20.0% and 25.0% solutions of poloxamer 407 (Kolliphor® P 407; P407) were applied. In all
of the investigated systems S:C ratio was 1:1 (w/w) and this value was kept constant through
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all experiments. The qualitative compositions of the investigated systems are presented in
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table 1. In order to obtain poloxamer 407 solutions, poloxamer 407 powder was dissolved in
cold deionized water (5.0±0.5°C). The cold solution was added to the mixture of oil,
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surfactant, and cosurfactant and the obtained mixtures were equilibrated at room temperature
before the visual inspection.

In the first step surfactant/cosurfactant mixture (Smix) was prepared and blended with the oil
phase in 1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2 and 9:1 weight ratio. Afterwards, the water phase

5
was added dropwise until the turbidity occurred. Moreover, the samples were inspected
visually for the increase of viscosity. Transparent liquids were classified as microemulsions.
The formulations D7-D10 (table 1) containing P407 solution were inspected visually at
ambient and 37.0±0.5°C in order to observe the possible gelation process.

2.3.Electrical conductivity studies

Electrical conductivity analyses were conducted for systems D3, D4 and D5. The tests
were performed along the dilution lines L1 – L6 corresponding to six different Smix : oil

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weight ratios (fig. 1). For all measurements, FiveEasy™ conductivity meter (Mettler Toledo,

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Greifensee, Switzerland) was used. The device was calibrated with 1413µS cm-1 and 12.88mS

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cm-1 standard solutions. Each measurement was done in triplicate and mean values were
calculated.

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2.4.Preparation of ME-based gels

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Systems D3, D4, and D5 (table 1) were selected for further investigations. The
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compositions of the selected microemulsions are presented in table 2.
The quantitative composition of MEs used to obtain semisolid gels is presented in table 2. In
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the first step, the appropriate amounts of Smix and oil phase were mixed in a beaker with the
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use of a magnetic stirrer. In the case of drug-loaded formulations, NPX was dissolved in the
obtained mixture. Afterwards, the water phase was added and the sample was stirred until the
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transparent system was obtained. In the next step, Carbopol® EZ-3 was spilled over the
sample surface and the system was sonicated at ultrasound frequency 47 000 Hz in three
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cycles, 180 seconds each. Next, the samples were mixed with a mechanical stirrer for 48 h at
700 rpm during the first 30 minutes and then at 300 rpm. In the case of placebo gels, the same
procedure without the drug dissolution step was applied. The compositions of gel samples
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are presented in table 3.

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2.5.Rheological analysis
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The rheological analysis was conducted at 25.0±0.5°C with HAAKE™ RheoStress™1

(ThermoScientific, Waltham, MA) rotational rheometer equipped with plate-plate geometry
(PP35Ti; diameter: 35 mm, gap: 1 mm) and temperature-controlled unit Thermo HAAKE™
DC 30. Before each measurement, the fresh sample was equilibrated on the plate for 2 min to
reach the desired temperature. All tests were performed in triplicate and mean values were
calculated.

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 2.5.1. Flow behavior analysis
Rotational experiments were conducted both in controlled shear rate (CR) and
controlled stress (CS) mode. In CR mode shear rate was increased linearly from 0 to 200.0 s -1
in 30 s and shear stress values were recorded. In CS mode the experiments were performed
under changing shear stress from 0 to 500.0 Pa in 15 s.

2.5.2. Oscillatory tests

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Two oscillatory tests were performed in this study. In the first one (the oscillatory

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stress sweep), the oscillatory stress was increased from 0.1 to 1000.0 Pa under the constant

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oscillatory frequency of 1.0 Hz. The obtained storage (G’) and loss (G”) moduli vs. stress
plots were used to estimate the linear viscoelasticity region and to select stress value of 3.0 Pa

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for the frequency sweep tests.
All oscillatory frequency sweep tests were carried out at a constant stress value, while the

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frequency values ranged from 0.1 to 100 Hz. The measured parameters were G’ and G”.
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2.6.Texture profile analysis
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For all measurements, TA-XT2i® (Stable Micro Systems) texture analyzer equipped
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with a steel cylindrical probe (diameter: 5 mm) was used. The velocity of the probe was set at
1 mm s-1. The samples (20.0 g) were placed in polypropylene containers and equilibrated at
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25.0±0.5°C for 15 minutes before the measurements. During the test, each sample was
compressed in two cycles with 20 s interval time and the applied force was monitored. The
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measurement depth was 18 mm. For each formulation, three independent measurements were
done with the use of fresh samples and the mean values were calculated. For all analyzed gels
hardness, adhesiveness, cohesiveness, gumminess, springiness and compressibility were
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assessed.
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2.7.Statistical analysis
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In the first step the obtained experimental data were analyzed with one-way
analysis of variance (ANOVA) to check if there are any statistically significant
differences between the investigated systems. Next, in order to answer the question in
which pairs of samples the statistically significant differences are observed, post hoc
Scheffe’s test based on multiple comparisons was performed. The mathematical details
related to both tests are described elsewhere [28]. The significance level in all experiments

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was set at 5%. The statistical calculations were performed with Statistica 12.5 software
(StatSoft Inc., Tulsa, OK).

3. Results and discussion

3.1.Pseudoternary phase diagrams
The obtained pseudoternary plots are presented in figures 2 and 3. The main factor
considered in this study was the extent of the area corresponding to isotropic liquid systems
classified as microemulsions. In the first step the impact of the oil phase on the monophasic

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area was investigated (fig. 2, D1-D3). The presented diagrams clearly indicate that systems

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with OA as an oil phase revealed better water solubilization potential which is most probably

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related to the structure and properties of the investigated oils. OA as an unsaturated fatty acid
reveals higher polarity than IPM and EO which are esters. Therefore, OA was selected as a

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component for further investigations. In the next step the impact of cosurfactant type on the
monophasic area extent was evaluated (fig. 2, D3-D6). It was shown that system with PG

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revealed significantly lower water solubilizing capacity while the results obtained for ethanol,
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isopropanol and Transcutol® P were similar. Taking into consideration these observations,
PG as a cosurfactant was excluded from the further studies.
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In the next step, the investigations concerning water phases were performed. The systems
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revealing the highest monophasic areas were selected for further analyses and deionized water
was replaced with poloxamer 407 solutions (fig. 3, D7-D10). The results depicted in
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pseudoternary plots are similar to those obtained for pure water systems. In this study
poloxamer 407 was selected as a potential gelling agent. According to the numerous literature
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reports, the micelles formed by its amphiphilic molecules can aggregate and form three-
dimensional structures [17,29–31]. This process is reversible and depends on the temperature
and the polymer concentration. Poloxamer 407 is considered as a valuable excipient in
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pharmaceutical technology. It was shown that it can be utilized to enhance the dissolution of
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the drugs revealing poor water solubility, improve the stability of particular active ingredients
and act as a gelling agent [17]. As a thermoresponsive polymer, it may be used for in-situ
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gelling formulations which are liquid at packing and storage temperature and form gels in the
physiological conditions [32,33]. However, the visual inspection of the samples D7-D9
containing 20.0% P407 solution instead of pure water showed no signs of gelation at the
increased temperature. The same results were obtained for D10 with 25.0% P407 solution as a
water phase. The observed properties may result from the presence of the oil phase, as well as
the cosurfactant which may be partially dissolved in the water phase. Therefore, as a

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thickening agent, Carbopol® EZ-3 was used because of its excellent compatibility with
microemulsions reported in numerous studies [34–38].

3.2.Electrical conductivity analysis

In this study, we focused on the conductivity changes observed during the titration of
oil / Smix mixture with water along six different dilution lines presented in the fig. 1. The aim
of these investigations was to gain an insight into the structure of analyzed systems and the

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relations between the particular phases. The conductivity curves plotted as a function of water

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content obtained in water titration experiments are presented in fig. 4.

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Electrical conductivity studies were performed in order to localize the transition points related
to the structural changes in the system and find the approximate boundaries of regions related

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to the occurrence of o/w, w/o and bicontinuous systems. Therefore, the conductivity was
monitored along L1-L6 dilution lines for the rapid changes occurring as a result of water

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addition. According to numerous literature reports [39–43], the observed changes are related
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to the structural transitions leading to different ME types. In the first stage of the experiment,
the water content is low and it is dispersed in a form of droplets in a continuous oil phase. Oil-
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continuous MEs reveal low conductivity values. The first sharp increase of conductivity
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during the increase of water content is related to the transition to the bicontinuous system,
with both polar and non-polar phases forming interpenetrating three-dimensional domains.
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Further addition of water results with an approximately linear increase of conductivity related
to the formation of water channels in ME structure. Finally, as a result of subsequent water
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addition, numerous water channels form a continuous phase and ME transforms into o/w type.
At this stage, the conductivity is either constant or drops. The conductivity measurements may
be used to study microstructural changes in MEs or to classify the particular system to one of
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three ME types. It is noteworthy that the structures of MEs are also important in terms of drug
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delivery. It was shown that depending on the structural features of the system different drug
release profiles may be obtained [44–46]. According to the results presented by Dojrdjevic et
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al. [45,46], o/w systems tended to show higher drug flux, while in the case of bicontinuous
ones the drug release was hampered. Similar investigations conducted by Podlogar et al. [47]
revealed that ketoprofen incorporated in different ME systems was released with zero-order
kinetic, however, o/w systems revealed higher drug flux than w/o ones. Therefore, assigning
the investigated ME system to one of three types is particularly important in terms of further
studies related to drug release and potential therapeutic effectiveness.

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In this study, three systems containing different cosurfactants have been investigated. In all of
them the point corresponding to the transition from w/o to bicontinuous ME was discernible
in the conductivity plots (fig. 4). However, the points related to the next transition, i.e. from
bicontinuous to o/w system, were observed only in some cases while in the other ones
transformation to turbid coarse emulsions occurred instead. It may be assumed that further
increase of water content in these systems led to a destabilization of bicontinuous MEs instead
of a transition to o/w systems.
In the experiments performed along the dilution lines L1-L3 for D3 systems containing

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ethanol, the first transition point was observed at about 20% water content. In the dilution

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lines L4-L6 the transition was observed at lower water concentrations, i.e. less than 5%.

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Similar results were obtained for D5 systems containing isopropanol. The transformation
from w/o to bicontinuous systems was observed at about 21-23% water content for the

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dilution lines L1-L3, while for L4-L6 at about 3-7%. Systems D4 containing Transcutol® as a
cosurfactant revealed transformation at about 13% for L1 and about 21% for L2. For the

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dilution lines L3-L6 the transition points were recorded at lower water concentrations (less
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than 10%).
The transition from bicontinuous to o/w MEs in D3 systems containing ethanol was
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discernible along the dilution lines L1-L5. In the experiment performed along the dilution
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line, L6 microemulsion transformed into coarse emulsion before reaching the second
transition point. In D5 systems containing isopropanol, the second transition point was
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recorded only in L1, L2 and L5 dilution lines. In D4 systems containing Transcutol® the
transformation into o/w MEs was observed only for L3-L5 lines. The approximate areas
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corresponding to the particular microemulsion types obtained in the described study are
depicted in phase diagrams in fig. 5. It is noteworthy that all three formulations selected for
further investigations were bicontinuous. The systems depicted in fig. 5 with red triangle have
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been used for gel preparation and the obtained semisolid products have been subjected to the
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mechanical studies involving rheological studies and texture profile analyses (TPA).
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3.3.Rheological analysis

3.3.1. Flow behavior analysis

All of the investigated gel samples revealed non-Newtonian, shear-thinning behavior.
The obtained flow curves (fig. 6) were fitted with the Casson model (Eq. 1) [48]. The

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calculated yield stress values along with the other rheological parameters are summarized in
table 3.

𝜏 0.5 = 𝜏𝐶0.5 + (𝜂𝐶 𝛾̇ )0.5 Eq. 1

where:
𝜏 – shear stress [Pa],
𝛾̇ – shear rate [s-1],

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𝜏𝐶 – Casson yield stress [Pa],

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𝜂𝐶 – Casson viscosity [Pa s].

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Strain vs. stress curves resulting from CS rotational experiments are depicted in fig. 7. The
results obtained in CR and CS mode, as well as in oscillatory stress sweep experiments are
consistent and reveal some tendencies in the mechanical behavior of the investigated samples.

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It was shown that the cosurfactant type had an impact on the rheological characteristics of the
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analyzed gels. Both in the placebo and NPX-loaded systems the highest yield stress values
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were obtained for the samples containing Transcutol® as a cosurfactant, while the lowest
values were recorded for gels with ethanol. The observed properties of the investigated
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samples correlate with the viscosity values of the cosurfactants (ethanol: 1.08 mPa s [49],
isopropanol: 2.05 mPa s [49] and Transcutol®: 3.85 mPa s [50]). This may indicate that the
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properties of the applied cosurfactant are the most important factor affecting the
characteristics of the final product.
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The impact of the drug on the flow behavior of the analyzed gels is not clear. In the study
performed in CR mode the samples containing isopropanol and Transcutol® as cosurfactants
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revealed the same behavior independent of the presence of the drug. In CS mode statistically
significant differences were observed for all samples.
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3.3.2. Oscillatory tests

The oscillatory tests were applied in order to investigate the viscoelastic behavior of
the ME-based gels. The results of the oscillatory stress sweep experiments are depicted in
fig. 8 (top). In this study storage (G’) and loss (G”) moduli were measured as a function of
changing oscillatory stress. The monitored parameters provide information on the structure
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and properties of the samples allowing also for the classification of the investigated systems.
The storage modulus is related to the elastic response of the sample, while the loss modulus
gives information on the part of the energy which is dissipated in a way typical for viscous
fluids. Another parameter derived from G’ and G” is loss tangent defined with eq. 2 [51]:

tan 𝛿 = 𝐺"/𝐺′ Eq. 2

where:

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tan 𝛿 – loss tangent [-]

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G” – loss modulus [Pa]

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G’ – storage modulus [Pa].

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The plots depicting tan δ vs. oscillatory stress and oscillatory frequency are presented
in fig. 9. Stress sweep test allowed for the estimation of the linear viscoelasticity range (LVR)

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which was the largest for the samples containing Transcutol®. In all investigated samples G’
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values in LVR prevailed over G”. Therefore, it may be assumed that the elastic properties of
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the analyzed systems prevail over the viscous ones. The oscillatory stress values recorded at
crossover points (G’=G”) are summarized in table 3 and correlated with yield stress values
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obtained in CR and CS modes. Statistical analysis performed for the obtained values revealed
that both cosurfactant type and the drug presence affected the characteristics of the sample.
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The results of frequency sweep tests indicate that in all analyzed samples both moduli
were only slightly dependent on the frequency values. In the initial frequency range up to 20.0
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Hz, the curves are approximately linear. Above about 20.0 Hz the increase of both G’ and G”
values is observed. The analyses of tan δ vs. frequency plots (fig. 9, bottom) indicate the
increase in tan δ values with the increasing frequency which may suggest the structural
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changes at higher frequencies. It may also be assumed that further increase of frequency
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values could result in gel structure damage. Moreover, for all gels storage modulus prevailed
over the loss modulus which is also reflected in the values of tan δ.
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Taking into consideration the chemical composition of the investigated systems it is obvious
that their three-dimensional structure is formed with non-covalent interactions between the
polymer chains. The viscoelasticity of the samples depends both on the composition of the
thickened liquid medium and on the presence of the drug. The results obtained in all

12
oscillatory tests indicate that NPX may affect the non-covalent interactions between the
adjacent carbopol chains decreasing the elasticity of the samples. Similar observation has
been made for microemulsion-based systems containing indomethacin [52]
The results of the oscillatory analyses allowed also for the classification of the
investigated systems. Even though there are numerous definitions of a gel available in the
literature, the strict explanation of this term seems to be difficult [53]. It is also important to
notice that the proposed definitions refer to different physicochemical or structural aspects of
the described systems. Similar conclusions can be made with respect to their classification.

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Taking into consideration the rheological approach presented by Clark and Ross-Murphy [54]

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gel classification can be made based on the results obtained in oscillatory studies and the

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observed relationships between storage and loss moduli. According to the proposed
terminology, strong gels can be characterized by the strongly pronounced prevalence of G'

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over G" and the independence of both parameters with respect to oscillatory frequency. In
weak gels called also structured liquids [53] G' is only a little higher than G" and both

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parameters show a slight dependency on the oscillatory frequency. In this classification
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system, two categories of polymer solutions are also specified, i.e. systems containing
entangled and non-entangled polymer chains. In the first category G' is lower than G" in the
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initial stage of the oscillatory study. During the test, both parameters increase and the
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crossover is observed. In the latter category G' is lower than G" in the whole experimental
range. In the analysis of the results obtained in the frequency sweep test, it is noteworthy that
ED

storage modulus is consistently higher that loss modulus which indicates the prevalence of
elastic properties over the viscous ones. However, in the whole experimental range, both
PT

moduli slightly increase in a linear manner which is typical for the systems falling into weak
gels category in the classification system presented by Clark and Ross-Murphy [54].
E
CC

3.4.Texture profile analysis

A

The texture profiles presented as force vs. time plots obtained for the investigated
samples are presented in fig. 10 while the parameters calculated with the use of texture
profiles are summarized in fig. 11. The statistical analysis revealed that all samples containing
ethanol and isopropanol did not differ significantly in terms of hardness, adhesiveness
gumminess and compressibility. In the case of Transcutol®-based formulations, higher values
for all of the mentioned parameters were recorded. It is important to notice that the presence

13
of the drug had no impact on the mechanical characteristics of the investigated systems. The
effects observed for cohesiveness parameter were similar, however, the differences between
the samples containing different cosurfactants were less distinct. Taking into consideration the
springiness parameter, the type of cosurfactant and the presence of NPX did not affect the
samples significantly.

Calixto et al. [55] emphasize the importance of hardness, compressibility and cohesiveness
for the properties of the marketed pharmaceutical product designed to be applied on the skin.

T
The information provided by texture profile analysis could be useful in terms of estimation of

IP
the sample behavior during the removal of the gel from the container and spreading at the

R
application site. Even though there are no absolute values of textural parameters that could be
used as predictors of the acceptability of a topical product, the results obtained in TPA can be

SC
applied to check the impact of particular factors on the mechanical behavior of the analyzed
samples. Taking into consideration the systems presented in this study it may be assumed that

U
the addition of NPX did not affect significantly the textural properties of the investigated
N
systems. Moreover, the type of cosurfactant seemed to be the most important factor in the
outcome of the study. It may also be assumed that the viscosity of the cosurfactant may play
A
an important role in the final performance of the topical semisolid product.
M

4. Conclusions
ED

In the presented study novel microemulsion-based polymer gels with poloxamer 182
(Synperonic®) as a surfactant were designed and characterized with special attention paid to
PT

the aspects important in terms of topical drug delivery. As a model active ingredient revealing
poor solubility in water naproxen, the non-steroidal anti-inflammatory drug was incorporated
in the investigated systems. The analysis of pseudoternary phase diagrams obtained with the
E

use of different oil phases and cosurfactants revealed that the extent of the area corresponding
CC

to transparent liquid systems depended mostly on the polarity of the oil phase. It was also
noted that better results were obtained for the systems containing monofunctional alcohols,
A

i.e. ethanol, isopropanol and Transcutol® than for propylene glycol containing two hydroxyl
moieties. Moreover, the potential of poloxamer 407 as a pharmaceutically accepted thickening
agent was investigated. However, no gelation was observed for systems containing both 20
and 25% polymer concentration.
The electrical conductivity measured as a function of water content in microemulsion systems
containing ethanol, isopropanol and Transcutol® as cosurfactants revealed typical changes

14
related to microstructural transitions observed as a result of polar phase addition. However, in
some cases, destabilization of ME and transformation into turbid coarse emulsion occurred
before o/w ME was formed. It was also revealed that all formulations selected for further
investigations were bicontinuous.
Flow behavior analyses performed in controlled shear rate and controlled shear stress modes
revealed that all semisolid formulations obtained with selected MEs and the polymer had
shear-thinning properties. This phenomenon can be considered as favorable both in terms of
application to the skin or mucous membranes and the design of industrial manufacturing

T
process. Taking into consideration the results of oscillatory tests, the investigated systems

IP
were classified as weak physical gels with non-covalent interactions between the polymer

R
chains and the other components of their three-dimensional structure. Yield stress values
obtained with different methods were consistent and depended on the cosurfactant type. It was

SC
noted that the properties of the final product corresponded to the viscosity of the cosurfactant.
It was also shown that as a result of drug presence yield stress values decreased which may be

U
attributed to the interaction of NPX molecules with polymer chains.
N
The texture profile analyses revealed that isopropanol- and ethanol-based systems did not
differ significantly in terms of mechanical properties. The samples containing Transcutol® as
A
a cosurfactant had higher hardness, adhesiveness, compressibility, gumminess and
M

springiness. It is also noteworthy that the presence of the drug did not affect the textural
properties of the analyzed semisolid formulations.
ED

5. Acknowledgements
PT

The project was financially supported by Poznan University of Medical Sciences grant No
502-01-03314429-03439. The authors would like to thank for Carbopol® EZ-3 samples
provided free of charge by Lubrizol, Synperonic™ PE/L 62 samples from Croda Poland and
E

naproxen samples kindly donated by EMO-FARM (Ksawerów, Poland). We would also like
CC

to thank Prof. Beata Czarnecka from the Department of Biomaterials and Experimental
Dentistry (Poznan University of Medical Sciences) for the access to the rheometer and Prof.
A

Janina Lulek from the Department of Pharmaceutical Technology (Poznan University of

Medical Sciences) for the scientific support.

15
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Figure captions

Fig. 1. Dilution lines followed in electrical conductivity study; symbols L1-L6 correspond to the different weight
ratios of oil and surfactant/cosurfactant mixtures (Smix).

T
R IP
SC
Fig. 2. Pseudoternary plots obtained for systems D1-D6.

U
N
A
M
ED
E PT
CC
A

21
 T
IP
R
SC
U
N
A
M
ED
E PT
CC
A

Fig. 3. Pseudoternary plots obtained for systems D7-D10.

22
 T
R IP
SC
U
N
A
M

Fig. 4. Conductivity plots obtained for D3, D4 and D5 systems along L1-L6 dilution lines.
ED
E PT
CC
A

23
 T
R IP
SC
U
N
A
M
ED
E PT
CC
A

Fig. 5. Pseudoternary plots for D3, D4 and D5 with depicted areas corresponding to w/o (red),
bicontinuous (labeled as bi; yellow) and o/w (blue) MEs. Grey areas were not investigated in the study.
The formulations used to obtain gels are depicted with red triangles.

24
 T
R IP
SC
U
N
A
M
ED
E PT
CC
A

Fig. 6. Flow curves obtained for placebo (left) and NPX-loaded (right) gels in CR mode.

25
 T
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Fig. 7. Flow curves obtained for placebo (left) and NPX-loaded (right) gels in CS mode.

R
SC
U
N
A
M
ED

Fig. 8. Oscillatory studies results obtained for placebo (left) and NPX-loaded (right) gels: stress sweep (top) and
frequency sweep (bottom) curves.
E PT
CC
A

26
 T
R IP
SC
U
N
Fig. 9. Tangent loss plots obtained in oscillatory stress sweep (top) and frequency sweep (bottom) tests for
placebo (left) and NPX-loaded (right) gels.
A
M
ED
E PT
CC
A

Fig. 10. Texture profiles obtained for placebo (left) and NPX-loaded gels (right).

27
 T
IP
Fig. 11. Texture profile parameters obtained for placebo and NPX-loaded gels with ethanol (black, E),
isopropanol (light grey, I) and Transcutol® (dark grey, T).

R
SC
U
N
A
M
ED
E PT
CC
A

28
 A
CC
EPT
ED
M
A
N
U
SC
RIP

29
T
Table captions

Table 1. Compositions of systems used to obtain pseudoternary phase diagrams. Surfactant (S) in all systems
was Synperonic™ PE/L 62 and S:C ratio in all systems was 1:1 (w/w).

Phase
diagram Water phase Oil phase Cosurfactant (C)
number

D1 Water EO Ethanol

D2 Water IPM Ethanol

T
IP
D3 Water OA Ethanol

R
D4 Water OA Transcutol® P

SC
D5 Water OA Isopropanol

D6 Water

Poloxamer 407
OA

U Propylene glycol
N
D7 solution (20.0%, OA Ethanol
w/w)
A
Poloxamer 407
D8 solution (20.0%, OA Transcutol® P
M

w/w)
Poloxamer 407
D9 solution (20.0%, OA Isopropanol
w/w)
ED

Poloxamer 407
D10 solution (25.0%, OA Transcutol® P
w/w)
E PT
CC
A

30
Table 2. The compositions of MEs. In all samples OA content was 5.0%, water content was 45.0% and
Synperonic® content was 25.0%.
Ethanol Transcutol® Isopropanol
Sample code
[%, w/w] [%, w/w] [%, w/w]
ME-E 25.0 - -
ME-T - 25.0 -
ME-I - - 25.0

Table 3. The compositions of ME-based gels. In all samples the concentration of Carbopol® EZ-3 was

T
3.0%.
ME-E ME-T ME-I NPX

IP
Sample code
[%, w/w] [%, w/w] [%, w/w] [%, w/w]
G-E 97.0 - - -
G-T - 97.0 - -

R
G-I - - 97.0 -
G-E-NPX 95.8 - - 1.2

SC
G-T-NPX - 95.8 - 1.2
G-I-NPX - - 95.8 1.2

U
Table 3. Rheological parameters obtained for placebo and NPX-loaded gels.
N
Yield stress [Pa]
Sample
A
CR CS SS
M

G-E 70.0±0.5 61.8±1.9 98.5±2.3

G-I 99.3±4.8 75.2±1.8 124.0±4.1

ED

G-T 161.6±7.3 118.8±1.2 306.13±2.20

PT

G-E-NPX 51.56±1.02 51.71±2.41 69.88±2.56

G-I-NPX 87.77±2.74 64.40±0.64 104.97±4.07

E

G-T-NPX 148.33±5.21 102.53±1.59 228.13±2.20

CC
A

31